Obesity affects more than one in three adults in the United States and contributes to the leading causes of morbidity and mortality. Addressing this public health emergency will require new approaches supported by deeper understanding of fundamental biological mechanisms. Fat cells or adipocytes have been a focus of basic research into obesity and metabolic disease. Mammals possess white adipocytes that are efficient at storing energy and thermogenic brown and beige adipocytes that can convert energy into heat. The activation of beige fat cells is associated with anti-obesity and anti-diabetes effects. Beige fat cell function is dependent on the transcriptional coregulatory protein PRDM16. While the molecular pathways governing beige adipocyte development and phenotype have been increasingly delineated, little is known about how beige adipocytes interact with other cell types within adipose tissue. We developed a novel three-dimensional whole tissue imaging technique to provide insights into cellular crosstalk within adipose tissue. We discovered a regional preference for beige fat biogenesis and a regulated interaction between beige adipocytes and the sympathetic nervous system (SNS). Employing mouse models with adipocyte-specific Prdm16 loss of function, we found that PRDM16 in adipocytes regulates sympathetic neurite projections to these cells. Here we propose to address two aims to elucidate the mechanisms underlying the crosstalk between beige adipocytes and the SNS. First, we will investigate whether sympathetic projections to adipose tissue are established during development and/or show plasticity in different physiological contexts. Second, we will test the role of several candidate pathways in modulating the communication between beige adipocytes and sympathetic neurons. In addition, we will employ a cell-type selective proteomic approach to comprehensively characterize the molecular components involved in the interaction between beige adipocytes and sympathetic projections. This proposal will provide an integrated view of adipose tissue, including its architecture and the interaction between beige adipocytes and projections from the sympathetic nervous system. These studies will produce important technical and conceptual advances that address knowledge gaps in our understanding of adipose tissue biology and will yield new mechanism-based therapeutic targets for the treatment of obesity and associated diseases.